Department of Neuropharmacology (D.P., F.S.), University of Tours,
Tours, France; and Solvay-Pharma (M.-O.C.), Suresnes, France
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Introduction |
The
antihypertensive properties of clonidine (Schmitt, 1977
), rilmenidine
(Sannajust and Head, 1994), and moxonidine (Ernsberger et al., 1993)
have been largely described, but very few authors determined the
cardiac antiarrhythmic properties of these compounds. These
centrally acting antihypertensive agents were originally considered as
preferential agonists of presynaptic
2-adrenoceptors and developed to inhibit the
activity of the sympathetic nervous system. In addition, it is well
established that the sympathetic nervous system plays an important role
in the genesis of certain types of arrhythmias (Manning and de van
Cotten, 1962; Hayashi et al., 1991). Therefore, imidazoline compounds,
which interact via imidazoline receptors (IRs) and
2-adrenoceptors with the activity of ortho-
and parasympathetic nervous systems, may present cardiac interests in therapeutics.
Since the first studies from Lathers et al. (1978) and Helke et al.
(1979), it has been clearly confirmed that the central nervous system
contributes to the antiarrhythmic side effects of numerous
antihypertensive agents such as 
-blockers and ganglionic blocking
agents, as well as 2-adrenoceptor agonists.
Subsequently, Thomas and Tripathi (1986), then Hayashi et al. (1991),
showed that 2-adrenoceptors were involved in
the central control of neurogenic arrhythmias and that hyperactivation
of the adrenergic tone is a triggering factor of several cardiac rhythm
disorders (e.g., ventricular, junctional, and atrial arrhythmias).
However, their mechanism of action, initially related to stimulation of central 2-adrenoceptors (Schmitt, 1977), was
brought under review by various studies (Bousquet et al., 1984;
Ernsberger et al., 1990), leading to the concept of the existence of
the new class of IRs. It has been suggested that IRs, mainly located in
the rostral ventrolateral medulla oblongata, are involved in the
central regulation of arterial blood pressure (BP), and that
stimulation of these receptors induces a sympathoinhibitory effect with
direct impact on the heart, kidneys, and vasculature (Göthert and
Molderings, 1992).
In the light of this concept of IR, we planned to examine the
antiarrhythmic properties of one of the most selective
I1 subtype of IR agonist, moxonidine. Its effects
have been previously studied in conscious Sprague-Dawley rats with
acute coronary occlusion without reperfusion, or when anesthetized,
with reperfusion (Lepran and Papp, 1994). These authors showed that a
preventive i.v. treatment with moxonidine significantly reduced both
ischemic and reperfusion arrhythmias. Furthermore, a recent study
reported potent antiarrhythmic effects of moxonidine on ouabain-induced
cardiac arrhythmias in guinea pigs (Mest et al., 1995).
The role of the central and autonomic nervous systems in cardiac
diseases has been clearly demonstrated in experimental and clinical
studies (Randall et al., 1976; Burch, 1978). Historically, the
hypothalamus has been recognized as one of the major structures involved in the cardiovascular responses induced by emotion and stress,
and observed during defense reaction in both animals and humans.
Electrical stimulation of various diencephalic structures has been
shown to induce extrasystoles (ES), ventricular tachycardias, and other
types of arrhythmias in rabbits (Evans, 1976; Zhou et al., 1994), cats
(Fuster and Weinberg, 1960; Manning and de van Cotten, 1962; Evans and
Gillis, 1978), dogs (Verrier et al., 1975), or rats (Morpurgo, 1968).
From such studies, we have chosen to electrically stimulate the
posterior hypothalamus of halothane-anesthetized rabbits to obtain a
reproducible experimental model of cardiac neurogenic arrhythmias.
Propranolol, a Vaughan-Williams (1984) class-II antiarrhythmic agent,
belongs to the series of 1/2-blockers. It exerts a membrane
stabilizing effect, by specifically increasing the electrical stability
of the myocardium, and is devoid of partial adrenoceptor agonist
properties. Moreover, Huang (1969) showed that propranolol (0.1-4
mg/kg i.v.), 5 min postinjection, prevents the development of cardiac
arrhythmias triggered by intrahypothalamic electrical stimulation in
anesthetized cat, and these effects can persist from ~20 to 30 min.
In Wistar rats, Elghozi et al. (1979) demonstrated that after i.v.
injection, propranolol diffuses into hypothalamic and medullary
structures. The retention by nuclei involved in BP and cardiac rate
regulation may be related to the antihypertensive and antiarrhythmic
actions via an inhibition of sympathetic tone. Furthermore, the C1 area
of the rostral ventrolateral medulla oblongata has been recognized as a
major site for the central hypotensive action of propranolol (Privitera
et al., 1988). As previously mentioned, apart from the direct cardiac
effects, the peripheral actions of propranolol-induced antihypertensive
and bradycardic effects may contribute to its antiarrhythmic
potency. Therefore, because propranolol appears to present
similar effects to those of moxonidine on sympathetic nerve activity,
we planned to investigate the antiarrhythmic effects of moxonidine in
halothane-anesthetized rabbits and compare them to those of propranolol.
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Materials and Methods |
Animals.
Six male normotensive Zika rabbits, aged 10 to 12 months and weighing 3.2 ± 0.3 kg, were used. They were obtained
from the Valteau colony (Valteau S.A., Bressuire, France) and were kept in our laboratory animal house for an acclimatization period of 8 to 12 days before use. The rabbits were maintained under standard conditions
of temperature (21 ± 1°C), lighting (12-h light/dark cycle;
lights on from 8:00 AM-8:00 PM; 100 ± 20 lux at cage level), humidity (60 ± 10%), and nonrecycled air, changed 15 to 20 times per hour. They received standard diet 112 (UAR Society,
Villemoisson-sur-Orge, France) containing less than 0.3% sodium, and
water ad libitum. The study was carried out in accordance with the
Guide for the Care and Use of Laboratory Animals presented by the
National Institutes of Health.
Surgical Preparation.
Under local anesthesia (2% xylocaine;
Astra-France Laboratories, Nanterre, France), a Teflon catheter (Jelco
22G, 25 mm; Johnson & Johnson Laboratories, New Brunswick, NJ) was
inserted into the ear medial artery, then connected to a pressure
transducer (RP 1500; Roucaire, Vélizy-Villacoublay, France) for
arterial BP measurement. A Teflon catheter (Jelco 24G, 19 mm; Johnson & Johnson Laboratories) was introduced into the ear marginal vein for
acute i.v. administration of substances.
After induction of anesthesia with propofol (Diprivan 1%, 10-20 mg/kg
i.v.; Zeneca-Pharma Laboratories, Cergy, France) and endotracheal cannulation (pediatric tube, 2.5 mm i.d.; Cole Foregger, New York, NY) each animal was connected to an inhalator (Fluotec Mark II; Cyprane, Keighley, UK), allowing spontaneous breathing of a
mixture (0.2-4%) of halothane (Fluothane; Zeneca-Pharma Laboratories) with room air, throughout the duration of electrode implantation and
experimentation. Body temperature was monitored and maintained at
38-39°C with a heating pad (Harvard homeothermic blanket control unit; Ealing S.A., Les Ulis, France).
Each rabbit was placed in a standard Horsley-Clarke stereotaxic device
(Société Réalisation-Application Mécanique,
Sartrouville, France) with a specific rabbit adaptor (Chatelier and
Buser, 1961). Then a 5-mm-diameter hole was made with a surgical drill
through the skull and a stainless steel, concentric, bipolar electrode (Rhodes Medical SNE 100 with connector, 21 mm length, 0.5 mm
o.d.; Phymep S.A., Paris, France) was inserted into the
posterior hypothalamus according to the following stereotaxic
coordinates related to the interaural line: A14.5 to A15.5; L0.5 to L1;
H0 to H +5 of Urban and Richard's atlas (1972) with the corrections
due to the adaptor (Poisson et al., 1974). The electrode was connected
to a power supply circuit delivering specific electrical stimulations. The implantation zone was determined as a function of the quality and
quantity of ECG disorders triggered by preliminary stimulations, and
then the electrode was subsequently solidarized to the cranial surface with three post screws (RRA-S1; Dentatus S.A., Hägersten, Sweden) and dental cement (Texton; SS-White, Gloucester, UK).
Three s.c. stainless steel ECG electrodes (23 gauge, 19 mm; Ets
Polylabo S.A., Strasbourg, France) were introduced under the skin of
forearms and hindlimbs of each animal for continuous ECG signals
acquisition from standard leads I, II, and III.
Experimental Protocol.
During each experiment, the electrode
of stimulation was connected to a main stimulator (Physiovar TR; Alvar,
Bordeaux, France) associated with two high-frequency isolation units.
The stimulator delivered two successive 1-ms rectangular pulses from
opposite polarity, with amplitude stimulation from 6 to 12 V and
frequency from 20 to 120 Hz. These latter parameters were varying in
function of delay between surgical implantation and experimental series and degree of anesthesia. This stimulator was programmed by another stimulator (Stimulator-T; Hugo Sachs Electronik, Hugstetten, Germany) which, by means of a time switch, delivered impulse trains for 30 s every 10, 20, or 45 min. The bipolar electrode was subsequently connected to two outputs of the two high-frequency links in
series with the center connected to the negative pole and the shaft to the positive pole. Stimulation was monitored by a scope (THS 720A; Tektronix S.A., Les Ulis, France) mounted in parallel with the circuit,
enabling accurate adjustment of voltage stimulation and maintenance of
a constant amplitude throughout the duration of experimentation.
Then, the arterial catheter was connected to a direct BP transducer and
systolic BP, diastolic BP, heart rate (HR), and the ECG biopotentials
were continuously recorded on a physiograph (Desk Model DMP-4B; NARCO,
Roucaire S.A., Vélizy-Villacoublay, France).
After a 20-min control period (C1) for which the hemodynamic and ECG
parameters were stable, the animals were subjected to three control
(S1, S2, S3) intrahypothalamic electrical stimulations (30-s duration
with a 10-min interval). After a 10-min recovery period, each animal
received an i.v. bolus injection of one of the three randomized
treatments (saline, moxonidine, propranolol). Fifteen minutes after
administration of the drug, four intrahypothalamic stimulations (S4,
S5, S6, S7) separated by 20 min were applied. Finally, to determine the
kinetics of antiarrhythmic effects of each treatment, S7 was followed
45 min later, by two additional postadministration stimulations (S8 and
S9) 20 min apart. A second control recording period (C2) was performed
to assess the reproducibility and stability of the experimental conditions.
On each experimental day, separated by a 2-week wash-out period, the
animals were subjected to arterial and venous ear catheterization. After propofol-induced anesthesia, the animals were connected via an
endotracheal cannula to the halothane-anesthesia maintenance system and
placed prone by using a foam. The preparative procedures for
stimulation and recording of hemodynamic and ECG parameters were
conducted as previously described.
At the end of the experimental series, each animal was sacrificed by an
overdose of sodium pentobarbital (Sanofi Santé Animale Laboratories, Libourne, France) and subjected to an intracarotid infusion of 200 ml of saline (NaCl 0.9%) followed by 150 ml of a 10%
formalin solution. Then, the implanted electrode was taken out of the
brain and the midbrain was removed for fixation in 10% formalin
before histological preparation for classic inclusion in paraffin. All
brains were cut in 10-µm sections to verify the intrahypothalamic
stimulating sites identified according to the stereotaxic landmarks
based on the atlas of Urban and Richard (1972) (see Fig.
1).
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Fig. 1.
Schematic diagrams of coronal brain sections close to
anterior planes A14.5, A15, and A15.5 (millimeters) related to the
interaural line according to the stereotaxic atlas of Urban and Richard
(1972) and demonstrating the distribution of stimulation sites ( ) in
six rabbits. CD, nucleus caudatus; HD, area hypothalamica dorsalis;
HDM, nucleus dorsomedialis hypothalami; HID, hippocampus dorsalis; HIV,
hippocampus ventralis; HP, area hypothalamica posterior; HVM, nucleus
ventromedialis hypothalami; HYL, area hypothalamica lateralis; RET,
nucleus reticularis thalami; cfx, columna fornicis; mt, tractus
mammillotegmentalis; to, tractus opticus.
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Drugs.
Moxonidine or
4-chloro-N-(4,5-dihydro-1H-imidazol-2-yl)-6-methoxy-2-methyl-5-pyrimidine,
kindly supplied by Solvay-Pharma (Suresnes, France), was administered
at the dose of 25 µg/kg i.v. Propranolol or
dl-(1-isopropylamino)-3-(-naphthyloxy)-2-propanol HCl was
purchased from Sigma Chemical Company (L'Isle d'Abeau, France) and
administered i.v. at a dose of 500 µg/kg.
Sterile saline (NaCl 0.9%), moxonidine (25 µg/kg), and propranolol
(500 µg/kg) were administered i.v. under a constant 250-µl volume
and followed by a 400-µl injection of sterile saline.
Data Analysis.
Systolic BP, diastolic BP, HR, and ECG
values continuously recorded on the chart physiograph were manually
measured off-line. The maximal values of mean arterial pressure (MAP)
and HR induced by each hypothalamic electrical stimulation (SMAP and
SHR, respectively) were determined at the peak value observed during
the 30-s stimulation and after the 2-min periods; they were then
compared with MAP and HR initial values (IMAP and IHR, respectively)
measured over the minute preceding each stimulation.
Arrhythmias of atrial or ventricular origins presented several kinds of
forms: isolated or by bursts of bigeminy, atrial or ventricular
tachycardias, torsades de pointes, auriculoventricular blocks, and the
QRS and T waves alterations (Fig. 2).
They were quantified by: 1) the number of extrasystoles (NbES) as
premature atrial or ventricular complexes; 2) the number of abnormal
ECG complexes (NbAC), consisting of ECG disorders of repolarization or
conduction type. Disorders of repolarization were based on reference
standard values reported by Kossakowski and Kuczynski (1973) in
anesthetized rabbits. In our experimental conditions, the values
observed in lead II were: magnitude of T wave: 0.1 ± 0.02 mV, QT
interval: 80 ± 6 ms and broad QRS interval: 40 ± 4 ms.
Abnormal values observed in magnitude of T wave (range: 0.17-0.3 mV),
QT interval (range: 100-160 ms), and broad QRS interval (range:
60-100 ms) were therefore considered as abnormal ECG complexes (ACs). In addition, the deeply inverted T waves, as well as cardiac auriculoventricular blocks have also been quantified as AC; 3) the
delay (d) of initiation of arrhythmias after the onset of hypothalamic
electrical stimulation and; 4) the total duration (D) of arrhythmias
for each period of stimulation.
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Fig. 2.
Representative traces showing the effects of
hypothalamic stimulation on arterial blood pressure (ABP) and ECG leads
II and III in one halothane-anesthetized rabbit in: control condition,
during the first stimulation (66 Hz, 7 V) (A); during the fourth
stimulation, 15 min after moxonidine (25 µg/kg i.v.) and 15 days
later in the same rabbit (B); during the third control stimulation (100 Hz, 10 V) (C); and during the fourth stimulation, 15 min after
propranolol (500 µg/kg i.v.) (D). Time scale indicates the delay
(seconds) after the onset of stimulation. A, typical ventricular ES and
two bursts of ventricular tachycardia [global parameters of
arrhythmias were 117 ES, 46 AC, onset delay of arrhythmia (d) = 8 s, duration of arrhythmia (D) = 36 s] associated to a fall
in ABP after an initial hypertension. B, antiarrhythmic action of
moxonidine characterized by a suppression of ventricular tachycardia,
decrease in NbES (22), NbAC (45), and arrhythmias duration (d = 12 s, D = 30 s). C, similar succession of various
ectopic beats (86 ES, 42 AC, d = 7 s, D = 39 s) to
(A). D, pronounced inhibition of arrhythmias after propranolol (0 ES, 2 AC, d = 16 s, D = 2 s).
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The hypotensive and bradycardic effects of moxonidine and propranolol
were measured during the first 15 min after injection and successively
during the minute preceding each intrahypothalamic stimulation (S4, S5,
S6, S7, S8, and S9).
Results are presented as means ± S.E. (n = 6 rabbits). For all parameters the effects of moxonidine were compared
with those of propranolol- and saline-injected groups, using a two-way
ANOVA for repeated measures. The significant differences between MAP and HR values measured before and during stimulation were determined for each treated group using Student's t test. In addition,
for every postinjection stimulation, we compared the effects of each treatment to the values measured before injection and to the effects of
other drugs, pairwise, by using the contrast method (SYSTAT software;
Evanston, IL), and P values less than .05 were considered significant.
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Results |
Anatomical Verification of Electrode Implantation Sites.
The
accuracy of intrahypothalamic stimulation sites was checked by
measuring the intensity and amplitude of the stimulation-induced arrhythmic responses and post-mortem histological verifications. Brain
sections of six rabbits showed three stimulation sites (see Fig. 1)
mainly located within the posterior (A14.5, L0.5, H +2 to H +3), dorsal
(A15, L0.5 to L1, H +1 to H +3) hypothalamic areas, and the dorsomedial
hypothalamic (A15.5, L0.5 to L0.8, H +1 to H +2) nucleus, according to
the atlas of Urban and Richard (1972).
Initial Cardiovascular Parameters.
In six
halothane-anesthetized rabbits, basal average MAP and HR values
determined during the minute preceding the three control stimulations
(S1, S2, S3) were 61 ± 1 mm Hg and 305 ± 4 beats/min, respectively (n = 54). There was no significant
difference in IMAP and IHR values between the three (moxonidine-,
propranolol-, and saline-) treated groups of animals.
Control intrahypothalamic stimulations (n = 54) induced
a significant (P < .001) increase (28%) in MAP
evaluated by the difference measured between SMAP and IMAP of +17 ± 2 mm Hg. The stimulation produced a significant (P < .001) tachycardic (20%) effect determined by the difference between
SHR and IHR (
HR = +60 ± 1 beats/min) (Fig. 2). The mean
values for control group are presented in Table 1. No significant difference during
control hypothalamic stimulations was observed between the three groups
of measured values (n = 18).
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TABLE 1
Effects of nine intrahypothalamic stimulations on cardiovascular and
arrhythmias parameters in six saline-treated anesthetized rabbits
Data are means ± S.E.
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Cardiovascular Effects of Substances.
The maximal hypotensive
and bradycardic effects of each substance were determined during the 15 min after i.v. bolus injection. Moxonidine (25 µg/kg) significantly
(P < .01) lowered MAP: 49 ± 5 versus 61 ± 4 mm Hg (MAP = 
12 ± 2 mm Hg) and was almost equihypotensive: 46 ± 4 versus 56 ± 4 mm Hg (MAP = 10 ± 2 mm Hg) to propranolol (500 µg/kg). In contrast,
propranolol produced a significant (P < .01) decrease
(25%) of resting HR: 234 ± 9 versus 312 ± 9 bpm
(HR = 78 ± 9 bpm). This effect was more pronounced after propranolol than moxonidine treatment (16%): 253 ± 8 versus 300 ± 15 beats/min (HR = 47 ± 10 beats/min) (Fig. 3).
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Fig. 3.
Effects of i.v. administration of moxonidine (MOX; 25 µg/kg), propranolol (PRO; 500 µg/kg), and saline (SAL) on MAP and
HR in six halothane-anesthetized rabbits. Filled columns represent mean
values (±S.E.) measured during the minute preceding injection; hatched
columns represent mean values measured during the 15 min after
injection. **P < .01, ***P < .001 versus before treatment;  P < .05, P < .01, P < .001 versus saline
treatment.
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Data in Fig. 4 indicate that 1 min before
each stimulation (S4, S5, S6, S7, S8, and S9) after injection, each
treatment significantly (P < .05) lowered MAP in
comparison with the mean IMAP recorded before each control stimulations
(S1, S2, and S3). Moxonidine exerted an hypotensive effect: 52 ± 5 versus 62 ± 3 mm Hg (IMAP = 15%), which became
significant (P < .05) only 2 h after injection whereas those of propranolol: 47 ± 5 versus 56 ± 3 mm Hg
(IMAP = 16%) appeared significant (P < .01)
from 55 to 75 min after the injection. The hypotensive action was
short-lived (20 min) for propranolol as well as for moxonidine.
Propranolol induced a significant (P < .01) decrease
in IHR: 219 ± 10 versus 309 ± 11 beats/min (IHR = 29%) compared with the mean IHR observed before each of the control
stimulations (S1, S2, S3) and to the saline-treated group: 300 ± 10 versus 307 ± 10 beats/min (HR = 2%) and moxonidine
treated-group: 280 ± 21 versus 299 ± 17 beats/min
(IHR = 6%). This decrease appeared 15 min postinjection and
persisted beyond the 140 min of recording.
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Fig. 4.
Effects of i.v. administration of moxonidine (,
MOX, 25 µg/kg), propranolol ( , PRO, 500 µg/kg), and saline ( ,
SAL) on IMAP and IHR before stimulations, or SMAP and SHR in six
anesthetized rabbits. Initial data represent mean values measured
during the minute preceding each stimulation (S1, S2, S3, S4, S5, S6,
S7, S8, and S9) and data under stimulation represent mean values
measured during the 30 s of each hypothalamic stimulation at the
maximum of variation. Time is indicated after the first stimulation
according to the protocol used, and dotted lines represent i.v.
injection (30 min after the start of the experiment). Vertical bars
show S.E. values. *P < .05, **P < .01, ***P < .001 versus
before propranolol injection; P < .05, P < .01, P < .001 versus before
moxonidine injection.
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The stimulation-induced hypertensive effect was significantly
(P < .01) lowered by moxonidine: SMAP = 67 ± 4 versus 75 ± 3 mm Hg (SMAP = 11%) from 35 to 55 min postinjection. Propranolol similarly prevented the SMAP increase:
69 ± 3 versus 77 ± 6 mm Hg (SMAP = 10%) but this
significant (P < .05) effect only appeared 35 min
postinjection. Moxonidine significantly (P < .05)
decreased SHR: 324 ± 9 versus 388 ± 28 beats/min
(SHR = 16%) 15 min after injection; this effect lasted 60 min. Fifteen minutes after injection, propranolol induced a significant
(P < .001) decrease in SHR: 230 ± 8 versus
351 ± 15 beats/min (SHR = 34%), which persisted throughout the duration of recording.
Effects of Control Intrahypothalamic Stimulations on ECG
Parameters.
The control intrahypothalamic stimulations (S1, S2,
and S3) induced for all animals cardiac arrhythmias that were mainly
characterized by sinusal or ventricular ES for whom mean number (NbES)
was 55 ± 2 (see Fig. 2 and Table 1 for control group). There was
no significant difference in NbES between each treated (saline,
moxonidine, propranolol) group of measured values (n = 18). Stimulations induced the emergence of NbAC characterized by
broadened QRS intervals, increased T wave, QT lengthening with
sometimes torsades de pointes, ST segment depression or elevation, and
auriculoventricular conduction disorders, eventually blocks. The mean
NbAC observed in animals was 37 ± 1 (Fig.
5). The average delay of arrhythmia onset
after poststimulation initiation observed in animals was 6.4 ± 0.4 s, and the mean duration of arrhythmia measured from the first
AC extended to 33.1 ± 1.4 s (Fig.
6). All parameters did not present any
significant difference between saline, moxonidine, and propranolol treatments.
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Fig. 5.
Effects of i.v. administration of moxonidine (,
MOX, 25 µg/kg), propranolol (, PRO, 500 µg/kg), and saline (,
SAL) on mean NbES and mean NbAC induced by posterior hypothalamic
stimulations in six anesthetized rabbits. Dotted line represents i.v.
injections (30 min after the start of experiment). Vertical bars show
S.E. values. *P < .05, **P < .01 versus before propranolol injection;
P < .05, P < .01, P < .001 versus before
moxonidine injection.
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Fig. 6.
Effects of i.v. administration of moxonidine (,
MOX, 25 µg/kg), propranolol (, PRO, 500 µg/kg), and saline (,
SAL) on delay (d) of onset and duration (D) of arrhythmias induced by
posterior hypothalamic stimulations in six anesthetized rabbits. Dotted
line represents i.v. injections (30 min after the start of the
experiment). Vertical bars show S.E. values. *P < .05, **P < .01 versus before propranolol
injection; P < .05, P < .01, P < .001 versus before
moxonidine injection.
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Effects of Administration of i.v. Drugs on Arrhythmias.
Moxonidine (25 µg/kg i.v.) induced a persistent (140 min after
injection) and significant (P < .01) decrease in NbES:
10 ± 4 versus 59 ± 6 (NbES = 83%) 15 min
postinjection (see Table 2 and Fig. 5).
Comparatively, propranolol (500 µg/kg i.v.) markedly (P < .01) decreased NbES: 2 ± 1 versus 53 ± 5 (NbES = 98%) triggered by stimulation in animals; this
effect began 15 min after injection and persisted until the end of the
experiment. Moxonidine induced a significant (P < .05)
decrease in NbAC: 28 ± 2 versus 37 ± 2 (NbAC = 33%) 75 min after i.v. injection. Propranolol significantly (P < .01) decreased the NbAC induced by stimulation:
14 ± 5 versus 37 ± 2 (NbAC = 79%) 15 min after
injection and this effect lasted throughout the duration of recording.
The effects of propranolol were significantly (P < .05) more pronounced (NbES = 15%; NbAC = 46%)
than those of moxonidine (Table 2). There were no significant effects
on NbES and NbAC in the saline-treated group (Table 1).
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TABLE 2
Comparative maximal antiarrhythmic effects of moxonidine (MOX, 25 µg/kg i.v.) and propranolol (PRO, 500 µg/kg i.v.) in six
halothane-anesthetized rabbits
Data are expressed in percentage of variation (%). Numbers in
parentheses indicate the kinetic of maximal antiarrhythmic effects
(time after i.v. injection of drug).
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Moxonidine treatment significantly (P < .01) prolonged
the average delay of onset of arrhythmia emergence as shown in Fig. 6
and Table 2. The maximal mean delay value was 12 ± 1 versus 7 ± 1 s (d = +65%). This effect started 15 min
postinjection and lasted 60 min. In contrast, propranolol did not
significantly increase the delay of arrhythmias: 13 ± 4 versus
6 ± 1 s (d = +188%). Fifteen to fifty-five min
after injection, moxonidine significantly (P < .01)
decreased the duration of arrhythmias: 22 ± 2 versus 34 ± 2 s (D = 35%). This effect was then attenuated but
became significant (P < .01; D = 23%) at
the end of the experiment (140 min). Propranolol significantly
(P < .01) reduced the duration of arrhythmias
triggered by stimulations: 15 ± 5 versus 35 ± 2 s
(D = 58%) but this effect, appearing 15 min after injection,
lasted only for 40 min. The comparison of moxonidine and propranolol
antiarrhythmic actions did not show any significant difference (Table
2), and saline injection did not modify the delay and duration of arrhythmias.
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Discussion |
The major finding of this study is that the most selective
I1 subtype of IR antihypertensive agent,
moxonidine, could not only prevent increases in MAP and HR induced by
electrical intrahypothalamic stimulation, but also exert potent
antiarrhythmic properties. These cardioprotective effects of moxonidine
have been demonstrated by using an halothane-anesthetized rabbit
arrhythmia model with posterior intrahypothalamic electrical
stimulations. Such conditions produced significant hypertensive and
tachycardic effects associated with neurogenic cardiac arrhythmias
predominantly characterized by premature and ectopic beats,
auriculoventricular blocks, and repolarization disorders.
First, we found that an i.v. dose of 25 µg/kg of moxonidine induced,
during the 15 min after the injection, significant but moderate
decreases in MAP and HR, which were similar to those observed after
administration of an i.v. dose of 500 µg/kg of propranolol (class II
-blocking agent of reference). Thus, if moxonidine and propranolol
were still equihypotensive 1 h after injection, propranolol
revealed to be more bradycardic (+9%) than moxonidine. Moreover,
propranolol, like moxonidine, inhibited the intrahypothalamic
stimulation-induced hypertension, and this effect lasted 1 h. The
two substances prevented the stimulation-induced tachycardia but
propranolol effect was longer (>2.5 h) than that of moxonidine (~1.5 h).
Second, moxonidine exhibited pronounced antiarrhythmic effects
characterized by profound decreases in NbES and NbAC, prolonged delay
of onset of arrhythmias, and important diminution of duration of
arrhythmias. These effects were slightly lower than those of propranolol, but lasted beyond the end of each experiment (140 min
after injection).
This study performed under long-lasting halothane anesthesia confirmed
the cardiodepressive and arrhythmogenic actions of this anesthetic
agent as previously reported by Maze and Smith (1983), Tranquilli et
al. (1986), and Hayashi et al. (1993) in anesthetized dogs. Halothane
lowered basal MAP (60 ± 1 versus 71 ± 4 mm Hg) and
accelerated HR levels (305 ± 4 versus 180 ± 10 beats/min)
in comparison with conscious rabbits (Sannajust and Head, 1994). In
addition, it is well established that halothane (as chloroform)
sensitizes the heart to arrhythmogenic effects of adrenaline, and this
property is currently used in experimental animal models of arrhythmias
(Caillard and Louis, 1980). Because the intrahypothalamic electrical
stimulations are responsible for an elevation in sympathetic tone and
adrenaline release from adrenal glands (Stoddard-Apter et al., 1983),
halothane potentiated the cardiac arrhythmias in our experimental
model. Moreover, it has been shown that the sympathetic nervous system
and adrenergic neuromediators (e.g., noradrenaline and adrenaline) are
directly involved in the BP and HR increases induced by posterior
intrahypothalamic stimulation in anesthetized and conscious cats
(Singewald and Philippu, 1996). However, the genesis of cardiac
arrhythmias of central origin may be related to a parasympathetic
nervous system activation as demonstrated by Manning and de van Cotten
(1962) in anesthetized cats. Similarly, the marked bradycardia obtained by Zhou et al. (1994) during electrical stimulation of lateral hypothalamus in isoflurane-anesthetized rabbits confirmed the important
role of cardiac vagal innervation in the genesis of arrhythmias.
The choice of moxonidine and propranolol doses used was based on
preliminary studies performed in our laboratory and by others (Lepran
and Papp, 1994) for which we observed that low doses (10-20 µg/kg
i.v.) of moxonidine induced slight antiarrhythmic effects, whereas a
higher dose (30 µg/kg i.v.) exerts pronounced and long-lasting bradycardic effects. Furthermore, we found that doses higher than 50 µg/kg i.v. could sometimes provoke respiratory or cardiac arrests in
some animals. The dose of propranolol (500 µg/kg) was chosen to be
equihypotensive with moxonidine.
In our experiments, the stimulation of specific posterior and dorsal
hypothalamic areas (as shown in Fig. 1) produced hypertension and
tachycardia, which suggest the involvement of the sympathetic nervous
system in the genesis of these neurogenic cardiac arrhythmias. This
sympathetic hyperactivity could be antagonized by central stimulation
of: 1) -adrenoceptors, as demonstrated by the effects of propranolol
in the anesthetized cat with hypothalamic electrical stimulation
(Huang, 1969); 2) 2-adrenoceptors, which are
stimulated by clonidine in the ouabain-induced guinea pig arrhythmia
model (Thomas and Tripathi, 1986); and 3) I1
subtype of IR predominantly stimulated by moxonidine in the same model
(Mest et al., 1995). In addition, the hypotensive action of these
compounds represents an additional benefit in the prevention of the
stimulation-induced BP increases that involve cardiac baroreflex
mechanisms responsible for arrhythmias (Evans and Gillis, 1978).
However, if the hypotensive effect of moxonidine in the conscious (Head
and Burke, 1991) and anesthetized rabbit has been initially attributed
to the stimulation of 2-adrenoceptors (Armah et al., 1988), additional studies demonstrated that the
sympathoinhibitory action of moxonidine (Ernsberger et al., 1993; Chan
et al., 1996) is more related to stimulation of
I1 subtype of IRs mainly located within the
rostral ventrolateral medulla oblongata (Haxhiu et al., 1994).
Moreover, Chan et al. (1996) showed that the i.v. moxonidine-induced
hypotension was antagonized with efaroxan (a mixed
I1-IRs and
2-adrenoceptor antagonist) given
intracisternally but not with 2-methoxyidazoxan (one of the most
selective 2-adrenoceptor antagonists).
Therefore, these findings suggest that the antihypertensive action of
moxonidine is preferentially mediated via stimulation of central
I1 subtype of IRs.
The antiarrhythmic effects of moxonidine observed in our study confirm
those previously reported by Lepran and Papp (1994). These authors
observed pronounced antiarrhythmic properties of moxonidine when
administered i.v. (10-100 µg/kg) during the acute phase of
experimental myocardial infarction to conscious coronary artery-ligated
rats and, when anesthetized, subjected to reperfusion arrhythmias. In
addition, Mest et al. (1995) demonstrated that moxonidine (100-400
µg/kg i.v.) dose dependently increased the threshold for
ouabain-induced arrhythmias in guinea pigs. They have shown that
moxonidine was 2-fold more effective than clonidine and as effective as
propranolol at a 10-fold higher dose. Our experiments show that
moxonidine at a relatively low dose (25 µg/kg i.v.) and propranolol
used at a 20-fold higher dose markedly decreased NbES and NbAC in
anesthetized rabbits, and confirm previous data reported by Mest et al.
(1995). In addition, these authors observed that the antiarrhythmic
action of moxonidine was completely prevented with an i.v. pretreatment
with efaroxan and was partially suppressed by a pretreatment with
idazoxan (a mixed ligand of I1 and
I2 subtypes of IR and an
2-adrenoceptor antagonist). Small doses (10 and 100 nM) of moxonidine and propranolol inhibited the appearance of
ES induced by application of aconitine in isolated preparations of
guinea pig atria. In contrast, a relatively high dose (1 µM) of
moxonidine was revealed to be ineffective in this model. Mest et al.
(1995) suggested that this biphasic action reflects a peripherally
mediated effect (only appearing at low doses) without any
arrhythmogenic effect (at high doses). Our experimental model of
neurogenic arrhythmias, like ouabain infusion-induced arrhythmia model,
is characterized by an elevation of sympathetic tone, which could be
antagonized by the central sympathoinhibitory action of moxonidine
preferentially mediated via stimulation of the I1
subtype of IR (Schäfer et al., 1995).
Furthermore, the chronic implantation of intrahypothalamic bipolar
electrodes in our normotensive rabbits allowed us to establish a within
animal statistical design, each animal being its own control. Such an
experimental model presents the advantages of performing long-lasting
experiments and determining the kinetics of action of drugs (such as
moxonidine and propranolol), which present prolonged antiarrhythmic
potency (see Table 2). Our results showed that the effects of
propranolol peaked 35 min postinjection and persisted for more than
2 h, except for the delay of onset and duration of arrhythmias (1 h only). Maximal antiarrhythmic effects of moxonidine were obtained 15 min after injection and, in some cases, exceeded the duration of the
experiments (140 min). We observed that the kinetics of the
antiarrhythmic action of moxonidine and propranolol was comparable
despite the difference in concentration. Likewise, the direct cardiac
and central effects, the propranolol-induced bradycardia, are known to
contribute to its antiarrhythmic properties. Thus, moxonidine
presents a slight bradycardic action at the dose used but no direct
cardiac effect, showing that its antiarrhythmic properties, as well as
those of propranolol, are probably of central origin (Mest et al.,
1995).
In conclusion, the results of this study demonstrate that moxonidine,
one of the most selective central I1 subtype of
IR antihypertensive agents, presents a dual therapeutic potency. A low
dose (25 µg/kg) of moxonidine inducing moderate hypotension and
pronounced bradycardia when administered i.v. to halothane-anesthetized
normotensive rabbits, can prevent neurogenic cardiac arrhythmias
resulting from repeated posterior hypothalamus electrical stimulations. The pronounced and long-lasting reductions in NbES, NbAC, and duration
of arrhythmias obtained after moxonidine were comparable to those
observed with propranolol treatment, a reference class II
antiarrhythmic agent. However, additional studies are required to
determine whether these antiarrhythmic effects are of central and/or
peripheral origin.
We thank E. Delaunay for assistance in programming and data
analysis and G. Chauveau for histological preparation of rabbit brains.
IR, imidazoline receptor;
AC, abnormal ECG
complex;
NbAC, number of abnormal ECG complexes;
BP, blood pressure;
ES, extrasystoles;
HR, heart rate;
IHR, initial heart rate;
IMAP, initial mean arterial pressure;
MAP, mean arterial pressure;
NbES, number of extrasystoles;
SHR, maximal heart rate under hypothalamic
stimulation;
SMAP, maximal mean arterial pressure under hypothalamic
stimulation.
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Abstract
Introduction
